Pancreatic ductal adenocarcinoma (PDAC) has the worst prognosis of any major malignancy with less than a 6% 5-year survival rate. It is one of the leading causes of cancer-related death in the developed world and is the fourth leading cause of cancer death in the United States. Patients with PDAC are often diagnosed at late stages with extensive local tumor invasion and early metastasis, presenting a major obstacle to all forms of therapy. Recently, cancer stem cells (CSCs) have come into focus as potential therapeutic targets in multiple cancers. The accumulation of mutations in CSCs enhances chemo/radiation resistance ablating the effect of therapy, leading to the cancer recurrence, characteristic of PDAC. Conversely, because CSCs are a unique subset of the tumor cell population, targeting these cells may lead to the identification of effective drugs for poorly treatable cancers such as PDAC. Epithelial-to-mesenchymal transition (EMT) is a well-documented characteristic of metastatic malignancies including PDAC. There is an increasing body of evidence linking EMT to a stem cell phenotype characterized by a capacity for self-renewal and differentiation, and expression of pluripotency factors. We recently demonstrated that doublecortin-like kinase 1 (DCLK1) is a putative pancreatic stem cell marker. DCLK1 is upregulated in several human solid tumor cancers, including colorectal, pancreatic, breast and prostate cancer. We also demonstrated that DCLK1 regulates multiple oncogenic and tumor supporting pathways and processes. Furthermore, siRNA mediated knockdown of DCLK1 results in growth arrest of pancreatic and colorectal cancer tumor xenografts. Moreover, introducing the KRASG12D mutation into Dclk1-positive cells in a Dclk1CreERT mouse model leads to rapid pancreatic cancer development synonymous to human PDAC. Recent studies also validated Dclk1 as a specific tumor stem cell marker in the pancreas and intestine. On the basis of these findings, our central hypothesis is that Dclk1 initiates and drives progression of PDAC and represents a viable therapeutic option worthy of investigation. We will test our hypotheses with the experiments proposed in the following three specific aims: Aim 1: To determine the functional and molecular mechanisms through which Dclk1+ cells initiate pancreatic tumorigenesis. Aim 2: To delineate the multi-compartmental role of Dclk1 deletion in pancreatic tumorigenesis. Aim 3: To demonstrate the feasibility of targeting Dclk1 in PDAC as a therapeutic approach. The current proposal includes innovative genetically engineered mouse models of PDAC, innovative cell lines, and innovative concepts that may lead to translational studies on PDAC.